We plan further elucidation and characterization of the regulatory processes of 5-HT biosynthesis in the brain using ions such as veratridine and K ion which depolarize the synaptosomes and increase the conversion of tryptophan to 5-HT. We will use C-1300 mouse neuroblastoma in which we have shown tryptophan to 5-HT conversion, as an in vitro model for enzyme, uptake, and conversion. We will further investigate the biochemical consequences of specific electrolytic lesions in the 5-HT system. Treatment with psychotomimetic amphetamines, shut down synaptosomal conversion of tryptophan to 5-HT without affecting tryptophan uptake. We intend to pursue these studies and to elucidate the mechanism of the shut down. We plan to further characterize the several aromatic amine N-methyltransferases in brain using the methyl donors 5-methyltetrahydrofolic acid and 5- adenosylmethionine and indoleamines and phenyethylamines as substrate acceptors. We will try to separate the enzymatically active protein factions by Sephedex columns and CaPO4 gels. We will extend substrate specificity studies of these enzymes to include aliphatic amines and imidazoleamines. If product quantities permit, Dr. Costa of NIMH has agreed to help us with cocrystallization and derivation using a double isotope technique. In addition, we plan to establish molecular weights of the enzymatic fractions, and to isolate and dialyze the inhibitor of the SAM-dependent enzyme.